This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Specific Aims and Hypothesis Prolonged exposure to crystalline silica in occupational and environmental settings induces chronic lung inflammation that can progress to fibrosis, i.e. silicosis. The interaction of pulmonary macrophages with silica is considered a key event in the disease process (1, 2). Furthermore, the environment of the lung is considered immunosuppressive;therefore, the induction of a chronic inflammatory condition like silicosis is the result of a disruption of the typical immunomodulatory environment of the lung. While a portion of this disruption is most likely due to the cytotoxic properties of silica particles, not all pulmonary macrophages die following interaction with the particulates. In fact we have documented increased activation of some macrophages in the mouse model. Respiratory exposure of mice (either Balb/c or C57Bl/6 common wild type strains) to crystalline silica results in pulmonary fibrosis, and has proven an effective model of the human disease. The Balb/c silicosis model has been shown, by our laboratory and others, to require Th2 immunity for progression to fibrosis (3-5). To date our laboratory has found several components associated with Th2 immunity to be significantly increased following exposure to crystalline silica including markers of alternatively activated macrophages (aaMac), interleukin (IL)-4+ interstitial cells, and insulin-like growth factor (IGF)-1. In addition, we have found a subset of pulmonary macrophages that express Akt, a key component of the survival pathway instigated by IGF-1, and appear to be predisposed to survival in the early stages of the disease. We propose to test the central hypothesis that multiple cells types and soluble factors that are associated with Th2 immunity are significantly increased in the Balb/c model of silicosis and that they all contribute to altered immune-activated environment of the disease. We will use the following two aims to test this hypothesis (as depicted in Fig. 1 schematic): Specific Aim 1: Evaluation of inflammasome activation in macrophage subsets of the lung following exposure to crystalline silica. Hypothesis: A subset of pulmonary macrophages induces the inflammasome pathway associated with IL-33 production and promotes a pro-fibrotic environment in the silicosis model. Using a combination of flow cytometry and mRNA analysis, pulmonary macrophage subsets will be defined based on inflammasome activation and IL-33 production. Specific Aim 2: Define the interaction of the inflammasome and survival (IGF-1-Akt) pathways following exposure to crystalline silica. Hypothesis: Silica induces both death and survival in particular macrophage subsets, both of which act synergistically to contribute to the pro-fibrotic environment of the silicosis model. Using in vitro culture systems (primary AM and bone marrow-derived macrophages) both the inflammasome and survival pathways will be activated with various stimulants in the presence of crystalline silica particles to assess their interaction in the development of a pro-fibrotic environment.